Phototransduction. The long-term objective is to understand the mechanisms underlying the electrical response to light in vertebrate photoreceptors. Single, isolated (solitary) rods and cones will be obtained after enzymatic treatment of the salamander retina. For cones, the influence of voltage, of calcium and, of cyclic nucleotides on the generator current will be determined under voltage-clamp conditions and compared with those already determined in rods. For rods, the effect on the light response of injecting antibodies directed against rod outer segments will be examined to generate specific reagents that can be used in analyzing the biochemical steps in the light response. Information on the underlying mechanisms is necessary in order to analyze and possibly correct pathological conditions that arise from toxicity and disease of photoreceptors. Rod-rod coupling. The precise role played by electrical coupling in shaping the light response within the rod cell layer needs to be determined. Pairs of rod cells will be used to measure the coupling conductance directly so that the effect of voltage and of levels of light adaptation can be accurately assessed. An analysis of coupling conductance will reveal its contribution to the time-course of electrical signals in the rod cell layer in situ. Cell-cell interaction. The long-term goal is to understand the types of cell-cell interactions that underlie the functional organization of the intact retina. The approach is to study the rules of synapse formation between identified cells in cultures of mature retinal neurons. The PI has developed a system whereby mature salamander neurons, grown on immobilized cell-surface antibody, will adhere to the coverslip, extend processes and reform synapses. This salamander system will be used to study the general question of synaptic specificitiy among mature CNS neurons. In order to examine specific questions in mammals, two separate systems using mature cat and monkey retinal cells will be developed drawing upon the experience gained with the salamander system. Cells will be identified by immunological, morphological and tracing methods, and synapse formation detected using intracellular recording techniques. Successful development of culture systems of mature mammalian neurons will be beneficial for clinical studies of regeneration in the damaged mature, mammalian central nervous system.